caa a22 4500 47711928X CHVBK 20180405111631.0 cr unu---uuuuu 170330e19960501xx s 000 0 eng 10.1007/BF00225844 doi (NATIONALLICENCE)springer-10.1007/BF00225844 The regulation of total creatine content in a myoblast cell line [Elektronische Daten] [Joseph Odoom, Graham Kemp, George Radda] Total cellular creatine content is an important bioenergetic parameter in skeletal muscle. To understand its regulation we investigated creatine transport and accumulation in the G8 cultured skeletal myoblast line. Like other cell types, these contain a creatine transporter, whose activity, measured using a radiolabelling technique, was saturable (Km = 110 ± 25 μM) and largely dependent on extracellular [Na+]. To study sustained influences on steady state creatine concentration we measured total cellular creatine content using a fluorimetric method in 48 h incubations. We found that the total cellular creatine content was relatively independent of extracellular creatine concentration, consistent with high affinity sodium-dependent uptake balanced by slow passive efflux. Accordingly, in creatine-free incubations net creatine efflux was slow ( 5 ± 1 % of basal creatine content per day over 6 days), while creatine content in 48 h incubations was reduced by 28 ± 13% of control by the Na+,K+-ATPase inhibitor ouabain. Creatine accumulation after 48 h was stimulated by treatment with the mixed α- and β-adrenergic agonist noradrenaline, the β-adrenergic agonist isoproterenol, the β2-agonist clenbuterol and the cAMP analogue N6,2′-O-dibutyryladenosine 3′,5′-cyclic monophosphate, but was unaffected by the α1 adrenergic agonist methoxamine. The noradrenaline enhancement of creatine accumulation at 48 h was inhibited by the mixed α- and β-antagonist labetalol and by the β-antagonist propranolol, but was unaffected by the α2 antagonist phentolamine; greater inhibition was caused by the β2 antagonist butoxamine than the β1 antagonist atenolol. Creatine accumulation at 48 h was increased to 230 ± 6% of control by insulin and by 140 ± 13% by IGF-I (both at 3 nM). Creatine accumulation at 48 h was also increased to 280 ± 40% of control by 3,3′,5-triiodothyronine (at 70 μM) and to 220 ± 35% of control by amylin (60 nM). As 3,3′,5-triiodothyronine, amylin and isoproterenol all stimulate the Na+,K+-ATPase, we suggest that they stimulate Na+-creatine cotransport indirectly by increasing the transmembrane [Na+] concentration gradient and membrane potential. Kluwer Academic Publishers, 1996 amylin nationallicence creatine nationallicence insulin-like growth factor 1 nationallicence Na+,K+-ATPase nationallicence triiodiothyronine nationallicence IGF-I : insulin-like growth factor I nationallicence IGF-II : insulin-like growth factor II nationallicence T3 : 3,3′,5-triiodothyronine nationallicence CGRP : calcitonin gene-related peptide nationallicence Odoom Joseph Department of Biochemistry, University of Oxford, Oxford, UK aut Kemp Graham Department of Biochemistry, University of Oxford, Oxford, UK aut Radda George Department of Biochemistry, University of Oxford, Oxford, UK aut Molecular and Cellular Biochemistry Kluwer Academic Publishers 158/2(1996-05-01), 179-188 0300-8177 158:2<179 1996 158 11010 https://doi.org/10.1007/BF00225844 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/BF00225844 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Odoom Joseph Department of Biochemistry, University of Oxford, Oxford, UK aut NATIONALLICENCE 700 1- Kemp Graham Department of Biochemistry, University of Oxford, Oxford, UK aut NATIONALLICENCE 700 1- Radda George Department of Biochemistry, University of Oxford, Oxford, UK aut NATIONALLICENCE 773 0- Molecular and Cellular Biochemistry Kluwer Academic Publishers 158/2(1996-05-01), 179-188 0300-8177 158:2<179 1996 158 11010 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer